Lipophilic but not hydrophilic statins selectively induce cell death in gynaecological cancers expressing high levels of HMGCoA reductase

Recent reports have suggested that statins induce cell death in certain epithelial cancers and that patients taking statins to reduce cholesterol levels possess lower cancer incidence. However, little is known about the mechanisms of action of different statins or the effects of these statins in gynaecological malignancies. The apoptotic potential of two lipophilic statins (lovastatin and simvastatin) and one hydrophilic statin (pravastatin) was assessed in cancer cell lines (ovarian, endometrial and cervical) and primary cultured cancerous and normal tissues. Cell viability was studied by MTS assays and apoptosis was confirmed by Western blotting of PARP and flow cytometry. The expressions of key apoptotic cascade proteins were analysed. Our results demonstrate that both lovastatin and simvastatin, but not pravastatin, selectively induced cell death in dose‐ and time‐dependent manner in ovarian, endometrial and cervical cancers. Little or no toxicity was observed with any statin on normal cells. Lipophilic statins induced activation of caspase‐8 and ‐9; BID cleavage, cytochrome C release and PARP cleavage. Statin‐sensitive cancers expressed high levels of HMG‐CoA reductase compared with resistant cultures. The effect of lipophilic statins was dependent on inhibition of enzymatic activity of HMG‐CoA reductase since mevalonate pre‐incubation almost completely abrogated the apoptotic effect. Moreover, the apoptotic effect involved the inhibition of synthesis of geranylgeranyl pyrophosphate rather than farnesyl pyrophosphate. In conclusion, lipophilic but not hydrophilic statins induce cell death through activation of extrinsic and intrinsic apoptotic cascades in cancerous cells from the human female genital tract, which express high levels of HMG‐CoA reductase. These results promote further investigation in the use of lipophilic statins as anticancer agents in gynaecological malignancies.     

Differential effects of simvastatin and pravastatin on expression of Alzheimer��s disease-related genes in human astrocytes and neuronal cells

Inhibitors of HMG-CoA reductase (statins) are widely used medications for reduction of cholesterol levels. Statin use significantly reduces risk of cardiovascular disease but has also been associated with lower risk of other diseases and conditions, including dementia. However, some reports suggest that statins also have detrimental effects on the brain. We provide evidence that simvastatin and pravastatin have significantly different effects on expression of genes related to neurodegeneration in astrocytes and neuroblastoma (SK-N-SH) cells in culture. Simvastatin significantly reduced expression of ABCA1 in astrocytes and neuroblastoma cells (by 79% and 97%, respectively; both P < 0.001). Pravastatin had a similar but attenuated effect on ABCA1 in astrocytes (−54%, P < 0.001) and neuroblastoma cells (−70%, P < 0.001). Simvastatin reduced expression of apolipoprotein E in astrocytes (P < 0.01). Furthermore, both statins reduced expression of microtubule-associated protein tau in astrocytes (P < 0.01), while both statins increased its expression in neuroblastoma cells (P < 0.01). In SK-N-SH cells, simvastatin significantly increased cyclin-dependent kinase 5 and glycogen synthase kinase 3β expression, while pravastatin increased amyloid precursor protein expression. Our data suggest that simvastatin and pravastatin differentially affect expression of genes involved in neurodegeneration and that statin-dependent gene expression regulation is cell type specific.—Dong, W., S. Vuletic, and J. J. Albers. Differential effects of simvastatin and pravastatin on expression of Alzheimer’s disease-related genes in human astrocytes and neuronal cells.     

Resveratrol for protection against statin toxicity in C2C12 and H9c2 cells

Statins are among the most commonly prescribed drugs for the treatment of high blood cholesterol. Myotoxicity of statins in certain individuals is often a severe side effect leading to withdrawal. Using C2C12 and H9c2 cells, both exhibiting characteristics of skeletal muscle cells, we addressed whether resveratrol (RSV) can prevent statin toxicity. Statins decreased cell viability in a dose and time‐dependent manner. Among the five statins tested, atorvastatin, simvastatin, lovastatin, pravastatin, and fluvastatin, simvastatin is the most toxic one. Simvastatin at 10 µM caused about 65% loss of metabolic activity as measured by 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide assays in C2C12 cells or H9c2 cells. Inhibition of metabolic activity correlates with an increase in caspase activity. RSV was found to protect H9c2 cells from simvastatin‐induced activation of caspase‐3/7. However, such protection was not found in C2C12 cells. This cell type‐dependent effect of RSV adds to the complexity in muscle cell toxicity of statins.     

Association between risk of myopathy and cholesterol-lowering effect: a comparison of all statins

In the present study, we examined the mechanisms underlying the cytotoxicity of pitavastatin, a new statin, and we compared the in vitro potencies of muscle cytotoxicity using a prototypic embryonal rhabdomyosarcoma cell line (RD cells), a typical side effect of statins and compared the cholesterol-lowering effects of statins using Hep G2 hepatoma cells. Pitavastatin reduced the number of viable cells and caused caspase-9 and -3/7 activation in a time- and concentration-dependent manner. The comparison of cytotoxities of statins showed that statins significantly reduced cell viability and markedly enhanced activity of caspase-3/7 in concentration-dependent manner. On the other hand, the effects of hydrophilic statins, pravastatin, rosuvastatin were very weak. The rank order of cytotoxicity was cerivastatin > simvastatin acid> fluvastatin > atorvastatin > lovastatin acid > pitavastatin > rosuvastatin, pravastatin. Statin-induced cytotoxicity is associated with these partition coefficients. On the other hand, the cholesterol-lowering effect of statins did not correlate with these partition coefficients and cytotoxicity. Thus, it is necessary to consider the association between risk of myopathy and cholesterol-lowering effect of a statin for precise use of statins.     

Hydroxymethylglutaryl--CoA reductase inhibitor inhibits induction of nitric oxide synthase in 3T3--L1 preadipocytes

Preadipocytes are considered to play a role in adipose tissue inflammation in obesity. The purpose of this study was to determine whether hydroxymethylglutaryl-CoA reductase inhibitor (statin) modulates the nitric oxide (NO) production via inducible NO synthase (iNOS) in preadipocytes. Undifferentiated 3T3-L1 cells, a model of preadipocytes, significantly produced NO by the treatment with the combination of lipopolysaccharide (L), tumor necrosis factor-alpha (T) and interferon-gamma (I). Pre-incubation with simvastatin, a lipophilic statin, or pravastatin, a hydrophilic one, dose-dependently inhibited the NO production in the LTI-treated cells. The effect of simvastatin was offset by mevalonate or geranylgeranyl pyrophosphate (GGPP) but not by squalene. The mRNA level for iNOS paralleled the NO production. The nuclear factor-kappaB (NF-kappaB) was activated by the LTI-treatment, and was inhibited by addition of simvastatin or pravastatin. Mevalonate or GGPP completely offset the effect of simvastatin. Simvastatin or pravastatin also decreased the LTI-stimulated interleukin-6 (IL-6) secretion. These effects of pravastatin were relatively weak compared with those of simvastatin. Y27632, an inhibitor of Rho kinase, also inhibited the LTI-induced NF-kappaB activation and iNOS expression, and decreased the production of NO and IL-6 in 3T3-L1 preadipocytes. These results suggest that statins, especially lipophilic types, inhibit induction of iNOS by inhibiting the small GTP-binding protein signal in preadipocytes.     

Attenuation of Acetylcholine Activated Potassium Current (IKACh) by Simvastatin,Not Pravastatin in Mouse Atrial Cardiomyocyte: Possible Atrial Fibrillation Preventing Effects of Statin

Statins, 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors, are associated with the prevention of atrial fibrillation (AF) by pleiotropic effects. Recent clinical trial studies have demonstrated conflicting results on anti-arrhythmia between lipophilic and hydrophilic statins. However, the underlying mechanisms responsible for anti-arrhythmogenic effects of statins are largely unexplored. In this study, we evaluated the different roles of lipophilic and hydrophilic statins (simvastatin and pravastatin, respectively) in acetylcholine (100 µM)-activated K⁺ current (I_KACh, recorded by nystatin-perforated whole cell patch clamp technique) which are important for AF initiation and maintenance in mouse atrial cardiomyocytes. Our results showed that simvastatin (1–10 µM) inhibited both peak and quasi-steady-state I_KACh in a dose-dependent manner. In contrast, pravastatin (10 µM) had no effect on I_KACh. Supplementation of substrates for the synthesis of cholesterol (mevalonate, geranylgeranyl pyrophosphate or farnesyl pyrophosphate) did not reverse the effect of simvastatin on I_KACh, suggesting a cholesterol-independent effect on I_KACh. Furthermore, supplementation of phosphatidylinositol 4,5-bisphosphate, extracellular perfusion of phospholipase C inhibitor or a protein kinase C (PKC) inhibitor had no effect on the inhibitory activity of simvastatin on I _KACh. Simvastatin also inhibits adenosine activated I_KACh, however, simvastatin does not inhibit I_KACh after activated by intracellular loading of GTP gamma S. Importantly, shortening of the action potential duration by acetylcholine was restored by simvastatin but not by pravastatin. Together, these findings demonstrate that lipophilic statins but not hydrophilic statins attenuate I_KACh in atrial cardiomyocytes via a mechanism that is independent of cholesterol synthesis or PKC pathway, but may be via the blockade of acetylcholine binding site. Our results may provide important background information for the use of statins in patients with AF.     

Simvastatin enhances induction of inducible nitric oxide synthase in 3T3-L1 adipocytes

The present study was designed to determine whether hydroxymethylglutaryl-CoA reductase inhibitors (statins) modulate the NO production via iNOS in adipocytes stimulated by lipopolysaccharide (L) and tumour necrosis factor-α (T). Well-differentiated 3T3-L1 adipocytes significantly produced NO by LT-treatment. Pre-incubation with simvastatin, a lipophilic statin, pravastatin, a hydrophilic one, or Y27632, an inhibitor of Rho kinase, further enhanced the production of NO. The effect of simvastatin was offset by mevalonate and geranylgeranyl pyrophosphate (GGPP) but not by squalene. The mRNA level for iNOS parallelled the NO production. The NF-κB was activated by the LT-treatment and was further enhanced by simvastatin, pravastatin or Y27632 addition. Mevalonate and GGPP completely offset the effect of simvastatin. Statins and Y27632 also further increased the interleukin-6 secretion in the LT-treated 3T3-L1 adipocytes. These results suggest that statins, especially lipophilic type, enhance induction of iNOS by inhibiting the small GTP-binding protein signal in adipocytes.     

The 3-hydroxy-3-methylglutaryl co-enzyme A reductase inhibitor pravastatin enhances neurite outgrowth in hippocampal neurons

Epidemiological studies demonstrate a relationship between statin [3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor] usage and reduced risk of developing Alzheimer's disease. To determine whether statins affect neuronal development, we treated cultured rat hippocampal neurons with pravastatin. After 4-48 h of treatment, pravastatin significantly increased the number of neurites produced by each cell and caused a corresponding increase in levels of the membrane phospholipid phosphatidylcholine. Pravastatin treatment also significantly increased neurite length and branching but did not affect cellular cholesterol levels. Co-incubation with mevalonate, but not cholesterol, abolished the stimulatory effect of pravastatin on neurite outgrowth. Treatment of neurons with isoprenoids also abolished the effect of pravastatin on neurite growth, suggesting that pravastatin may stimulate neuritogenesis by preventing isoprenylation of signaling molecules such as the Rho family of small GTPases. A specific inhibitor of geranylgeranylation, but not farnesylation, mimicked the stimulatory effect of pravastatin on neuritogenesis. Pravastatin treatment significantly decreased levels of membrane-associated RhoA. These data suggest that pravastatin treatment increases neurite outgrowth and may do so via inhibiting the activity of geranylgeranylated proteins such as RhoA.     

Coenzyme Q10 protects against β-cell toxicity induced by pravastatin treatment of hypercholesterolemia

New onset of diabetes is associated with the use of statins. We have recently demonstrated that pravastatin-treated hypercholesterolemic LDL receptor knockout (LDLr^−/−) mice exhibit reductions in insulin secretion and increased islet cell death and oxidative stress. Here, we hypothesized that these diabetogenic effects of pravastatin could be counteracted by treatment with the antioxidant coenzyme Q ₁₀ (CoQ ₁₀), an intermediate generated in the cholesterol synthesis pathway. LDLr ^−/− mice were treated with pravastatin and/or CoQ ₁₀ for 2 months. Pravastatin treatment resulted in a 75% decrease of liver CoQ ₁₀ content. Dietary CoQ ₁₀ supplementation of pravastatin-treated mice reversed fasting hyperglycemia, improved glucose tolerance (20%) and insulin sensitivity (>2-fold), and fully restored islet glucose-stimulated insulin secretion impaired by pravastatin (40%). Pravastatin had no effect on insulin secretion of wild-type mice. In vitro, insulin-secreting INS1E cells cotreated with CoQ ₁₀ were protected from cell death and oxidative stress induced by pravastatin. Simvastatin and atorvastatin were more potent in inducing dose-dependent INS1E cell death (10–15-fold), which were also attenuated by CoQ ₁₀ cotreatment. Together, these results demonstrate that statins impair β-cell redox balance, function and viability. However, CoQ ₁₀ supplementation can protect the statins detrimental effects on the endocrine pancreas.     

Statins inhibit growth of human endometrial stromal cells independently of cholesterol availability

Endometriosis is characterized by ectopic growth of endometrial tissues. Statins, inhibitors of 3-hydroxy-3methylglutaryl-coenzyme A reductase (HMGCR), have been shown to decrease proliferation of several mesenchymal tissues. Actions of statins may be related to decreased availability of cholesterol as well as intermediate metabolites of the mevalonate pathway downstream of HMGCR. This study was designed to evaluate effects of statins on growth of endometrial stromal cells and to investigate mechanisms of these effects. Human endometrial stromal cells were cultured in the absence and in the presence of serum and with or without mevastatin and simvastatin. DNA synthesis and viable cell numbers were determined. Effects of statins were also evaluated in the presence of mevalonate and squalene. Furthermore, effects on phosphorylation of mitogen-activated protein kinase 3/1 (MAPK3/1) (also known as extracellular signal-regulated kinase [ERK1/2]) were determined. Mevastatin and simvastatin induced a concentration-dependent inhibition of DNA synthesis and viable cell count in chemically defined media and in the presence of serum. Mevalonate, but not squalene, abrogated inhibitory effects of statins on cell proliferation. Statins inhibited MAPK3/1 phosphorylation. This is the first study demonstrating that statins inhibit growth of endometrial stromal cells. This effect is also demonstrable in the presence of a supply of cholesterol and may be related to decreased activation of MAPK3/1. The present observations may be relevant to potential therapeutic use of statins in conditions such as endometriosis.     

High doses of simvastatin, pravastatin, and cholesterol reduce brain cholesterol synthesis in guinea pigs

Recent epidemiological studies suggest that inhibitors of 3-hydroxy-3-methyl-glutaryl CoA reductase, so-called statins, are effective in lowering the prevalence of Alzheimer's disease. Whether the effect of statins is due to a local inhibition of cholesterol synthesis in the brain or whether it is mediated by the reduced levels of cholesterol in the circulation is not known. In the present work, we tested the possibility that high doses of lipophilic and hydrophilic statins, simvastatin and pravastatin, respectively, or a diet high in cholesterol could affect cholesterol homeostasis in the brain of guinea pigs. The total brain cholesterol levels were not affected by high-dose simvastatin or pravastatin treatment. Significantly lower levels of the cholesterol precursor lathosterol and its ratio to cholesterol were found in the brains of simvastatin and pravastatin-treated animals. 24S-Hydroxycholesterol, the transportable form of cholesterol across the blood-brain barrier, was significantly lower in the brain of pravastatin-treated animals. Excessive cholesterol feeding resulted in higher serum cholesterol levels but did not affect total brain cholesterol level. However, de novo cholesterol synthesis in the brain seemed to be down-regulated, as indicated by lower absolute levels and cholesterol-related ratios of lathosterol compared with controls. The passage of deuterium-labeled cholesterol across the blood-brain barrier in one animal was found to be approximately 1%. Our results suggest that brain cholesterol synthesis in guinea pigs can be slightly, but significantly, influenced by high doses of lipophilic and hydrophilic statins as well as by high dietary cholesterol intake, while total brain cholesterol content and thus, cholesterol homeostasis is maintained.     

Effects of simvastatin and pravastatin on peroxidation of erythrocyte plasma membrane lipids in patients with type 2 hypercholesterolemia

Since hypercholesterolemia directly modifies the composition of erythrocytes plasma membrane, the influence of statins on erythrocytes has been researched. The beneficial effects of statins on clinical events may involve mechanisms that modify endothelial dysfunction, plaque stability, thrombus formation and inflammatory responses. The aim of the study was to evaluate the hypolipemic efficacy and effects of pravastatin and simvastatin on erythrocyte membrane fluidity and damage of erythrocytes in patients with type 2 hypercholesterolemia in comparison with a control group of healthy subjects. The study involved 53 patients affected by type 2 hypercholesterolemia (mean age, 53.3 +/- 10.3) with initial total serum cholesterol (TC) levels > 250 mg/dL, LDL-cholesterol (LDL-C) levels > 170 mg/dL, and triglycerides (TG) levels < 400 mg/dL. The control group consisted of 30 healthy individuals (mean age 56.9 +/- 6.3). Statins were given for 12 weeks. The dosages for oral administration of simvastatin and pravastatin were 20 mg/day. Laboratory tests were carried out before and after 4 and 12 weeks of the pharmacological treatment. The damage to plasma membrane of erythrocytes was measured on the basis of lipid peroxidation. The fluidity of plasma membrane of erythrocytes was determined by electron paramagnetic resonance (EPR) spectroscopy, using two spin labels: 5-DSA and 16-DSA. The cholesterol level in the membrane of red blood cells was estimated. Simvastatin and pravastatin reduced the total cholesterol concentration and LDL-cholesterol in plasma, as well as the cholesterol concentration in erythrocytes membranes. Hypercholesterolemia induced changes in the basic properties of human erythrocyte plasma membrane, including its fluidity and the intensity of lipid peroxidation. These results indicate that the simvastatin and pravastatin therapy reverses the alteration in the erythrocyte plasma membrane properties.     

Simvastatin enhances induction of inducible nitric oxide synthase in 3T3-L1 adipocytes

The present study was designed to determine whether hydroxymethylglutaryl-CoA reductase inhibitors (statins) modulate the NO production via iNOS in adipocytes stimulated by lipopolysaccharide (L) and tumour necrosis factor-alpha (T). Well-differentiated 3T3-L1 adipocytes significantly produced NO by LT-treatment. Pre-incubation with simvastatin, a lipophilic statin, pravastatin, a hydrophilic one, or Y27632, an inhibitor of Rho kinase, further enhanced the production of NO. The effect of simvastatin was offset by mevalonate and geranylgeranyl pyrophosphate (GGPP) but not by squalene. The mRNA level for iNOS parallelled the NO production. The NF-kappaB was activated by the LT-treatment and was further enhanced by simvastatin, pravastatin or Y27632 addition. Mevalonate and GGPP completely offset the effect of simvastatin. Statins and Y27632 also further increased the interleukin-6 secretion in the LT-treated 3T3-L1 adipocytes. These results suggest that statins, especially lipophilic type, enhance induction of iNOS by inhibiting the small GTP-binding protein signal in adipocytes.     

Statins enhance clonal growth of late outgrowth endothelial progenitors and increase myocardial capillary density in the chronically ischemic heart

Background

Coronary artery disease and ischemic heart disease are leading causes of heart failure and death. Reduced blood flow to heart tissue leads to decreased heart function and symptoms of heart failure. Therapies to improve heart function in chronic coronary artery disease are important to identify. HMG-CoA reductase inhibitors (statins) are an important therapy for prevention of coronary artery disease, but also have non-cholesterol lowering effects. Our prior work showed that pravastatin improves contractile function in the chronically ischemic heart in pigs. Endothelial progenitor cells are a potential source of new blood vessels in ischemic tissues. While statins are known to increase the number of early outgrowth endothelial progenitor cells, their effects on late outgrowth endothelial progenitor cells (LOEPCs) and capillary density in ischemic heart tissue are not known. We hypothesized that statins exert positive effects on the mobilization and growth of late outgrowth EPCs, and capillary density in ischemic heart tissue.

Methodology/Principal Findings

We determined the effects of statins on the mobilization and growth of late outgrowth endothelial progenitor cells from pigs. We also determined the density of capillaries in myocardial tissue in pigs with chronic myocardial ischemia with or without treatment with pravastatin. Pravastatin therapy resulted in greater than two-fold increase in CD31+ LOEPCs versus untreated animals. Addition of pravastatin or simvastatin to blood mononuclear cells increased the number of LOEPCs greater than three fold in culture. Finally, in animals with chronic myocardial ischemia, pravastatin increased capillary density 46%.

Conclusions

Statins promote the derivation, mobilization, and clonal growth of LOEPCs. Pravastatin therapy in vivo increases myocardial capillary density in chronically ischemic myocardium, providing an in vivo correlate for the effects of statins on LOEPC growth in vitro. Our findings provide evidence that statin therapy can increase the density of capillaries in the chronically ischemic heart.     

The importance of LDL and cholesterol metabolism for prostate epithelial cell growth

Cholesterol-lowering treatment has been suggested to delay progression of prostate cancer by decreasing serum LDL. We studied in vitro the effect of extracellular LDL-cholesterol on the number of prostate epithelial cells and on the expression of key regulators of cholesterol metabolism. Two normal prostatic epithelial cell lines (P96E, P97E), two in vitro immortalized epithelial cell lines (PWR-1E, RWPE-1) and two cancer cell lines (LNCaP and VCaP) were grown in cholesterol-deficient conditions. Cells were treated with 1-50 μg/ml LDL-cholesterol and/or 100 nM simvastatin for seven days. Cell number relative to control was measured with crystal violet staining. Changes in mRNA and protein expression of key effectors in cholesterol metabolism (HMGCR, LDLR, SREBP2 and ABCA1) were measured with RT-PCR and immunoblotting, respectively. LDL increased the relative cell number of prostate cancer cell lines, but reduced the number of normal epithelial cells at high concentrations. Treatment with cholesterol-lowering simvastatin induced up to 90% reduction in relative cell number of normal cell lines but a 15-20% reduction in relative number of cancer cells, an effect accompanied by sharp upregulation of HMGCR and LDLR. These effects were prevented by LDL. Compared to the normal cells, prostate cancer cells showed high expression of cholesterol-producing HMGCR but failed to express the major cholesterol exporter ABCA1. LDL increased relative cell number of cancer cell lines, and these cells were less vulnerable than normal cells to cholesterol-lowering simvastatin treatment. Our study supports the importance of LDL for prostate cancer cells, and suggests that cholesterol metabolism in prostate cancer has been reprogrammed to increased production in order to support rapid cell growth.     

Simvastatin promotes osteoblast differentiation and mineralization in MC3T3-E1 cells

The cholesterol-lowering drug, simvastatin, is a pro-drug of a potent 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitor and inhibits cholesterol synthesis in humans and animals. In addition, the bone effects of statins including simvastatin are being studied. We assessed the effects of simvastatin on osteoblastic differentiation in nontransformed osteoblastic cells (MC3T3-E1) and rat bone marrow cells. Simvastatin enhanced alkaline phosphatase (ALP) activity and mineralization in a dose- and time-dependent fashion. This stimulatory effect of the statin was observed at relatively low doses (significant at 10(-8) M and maximal at 10(-7) M). Northern blot analysis showed that the statin (10(-7) M) increased in bone morphogenetic protein-2 as well as ALP mRNA concentrations in MC3T3-E1 cells. Simvastatin (10(-7) M) slightly increased in type I collagen mRNA abundance throughout the culture period, whereas it markedly inhibited the gene expression of collagenase-1 between days 14 and 22 of culture. These results indicate that simvastatin has anabolic effects on bone through the promotion of osteoblastic differentiation, suggesting that it could be used for the treatment of common metabolic bone diseases such as osteoporosis.     

Effects of statins on myocardial and coronary artery response to ischemia-reperfusion

This study tested the hypotheses that (i) lipophilic statins (atorvastatin and simvastatin) impair ventricular recovery from myocardial ischemia-reperfusion, owing to their greater myocyte permeability, compared with a hydrophilic statin (pravastatin), and (ii) statins enhance endothelium-dependent vasodilation of isolated coronary arteries from the ischemic region. Farm pigs consumed chow supplemented with atorvastatin (2.5 mg.kg(-1).d(-1); n=6), pravastatin (10 (n=3) or 20 (n=2) mg.kg(-1).d(-1)), simvastatin (5 mg.kg(-1).d(-1); n=6), or no statin (control; n=6) for 3 weeks. Animals were anesthetized and instrumented to measure regional (% segment shortening) and global (dP/dt max) left ventricular (LV) function during coronary artery occlusion (10 min) and reperfusion (30 min). Coronary resistance (i.d. = 119 +/- 3 microm) and conductance (i.d. = 487 +/- 11 microm) arteries were isolated from the ischemic region to measure receptor-dependent (acetylcholine (ACh)) and -independent (KCl) vasoconstriction, and endothelium-dependent (bradykinin (BK)) and -independent (sodium nitroprusside (SNP)) vasodilation. At 30 min reperfusion, neither percent recovery of regional ventricular function (atorvastatin, 24% +/- 15%; pravastatin, 36% +/- 13%; simvastatin, 29% +/- 13%; control, 36% +/- 13%) nor percent recovery of global LV cardiac function differed among groups. However, BK-induced vasorelaxation of coronary conductance vessels was greater (P<0.05) in statins versus controls, and ACh-induced vasoconstriction was less in simvastatin-treated animals, suggesting the potential for enhanced coronary arterial blood flow to the jeopardized region. In conclusion, our data suggest that ischemia-induced myocardial stunning is similar among pigs treated for 3 weeks with atorvastatin, pravastatin, or simvastatin, even though statin treatment appears to augment endothelium-dependent vasodilation of conductance, but not resistance, vessels subjected to ischemia-reperfusion.     

Cholesterol-Secreting and Statin-Responsive Hepatocytes from Human ES and iPS Cells to Model Hepatic Involvement in Cardiovascular Health

Hepatocytes play a central and crucial role in cholesterol and lipid homeostasis, and their proper function is of key importance for cardiovascular health. In particular, hepatocytes (especially periportal hepatocytes) endogenously synthesize large amounts of cholesterol and secrete it into circulating blood via apolipoprotein particles. Cholesterol-secreting hepatocytes are also the clinically-relevant cells targeted by statin treatment in vivo. The study of cholesterol homeostasis is largely restricted to the use of animal models and immortalized cell lines that do not recapitulate those key aspects of normal human hepatocyte function that result from genetic variation of individuals within a population. Hepatocyte-like cells (HLCs) derived from human embryonic and induced pluripotent stem cells can provide a cell culture model for the study of cholesterol homeostasis, dyslipidemias, the action of statins and other pharmaceuticals important for cardiovascular health. We have analyzed expression of core components for cholesterol homeostasis in untreated human iPS cells and in response to pravastatin. Here we show the production of differentiated cells resembling periportal hepatocytes from human pluripotent stem cells. These cells express a broad range of apolipoproteins required for secretion and elimination of serum cholesterol, actively secrete cholesterol into the medium, and respond functionally to statin treatment by reduced cholesterol secretion. Our research shows that HLCs derived from human pluripotent cells provide a robust cell culture system for the investigation of the hepatic contribution to human cholesterol homeostasis at both cellular and molecular levels. Importantly, it permits for the first time to also functionally assess the impact of genetic polymorphisms on cholesterol homeostasis. Finally, the system will also be useful for mechanistic studies of heritable dyslipidemias, drug discovery, and investigation of modes of action of cholesterol-modulatory drugs.     

Statin induces inhibition of triple negative breast cancer (TNBC) cells via PI3K pathway

Primary TNBCs are treated as if they were a single disease entity, yet it is clear they do not behave as a single entity in response to current therapies. Recently, we reported that statins might have a potential benefit for TNBCs associated with ets-1 overexpression. The aim of this study is to investigate the role of PTEN loss in the effects of statin on TNBC cells. In addition, we analyze the relationship between AKT downstream pathways and the effects of statin on TNBC cells. We investigated the effect of a statin on TNBC cells and analyzed the association of PI3K pathways using various TNBC cells in terms of PTEN loss and AKT pathways. Simvastatin treatments resulted in decreased cell viabilities in various TNBC cell lines. Compared with PTEN wild-type TNBC cells, PTEN mutant-type TNBC cells showed a decreased response to simvastatin. Expressions of phosphorylated Akt and total Akt showed an inverse relationship with PTEN expression. The TNBC cell lines, which showed increased expression of p-Akt, appeared to attenuate the expression of p-Akt by PTEN loss in simvastatin-treated TNBC cells. The Akt inhibitor, LY294002, augmented the effect of simvastatin on PTEN wild-type TNBC cells. Simvastatin induces inhibition of TNBC cells via PI3K pathway activation.